Base excision repair (BER) is a primary mechanism for maintaining genome integrity. A large variety of modified nucleotides resulting from DNA oxidation and alkylation are removed by glycosylases (Friedberg et al., 2006). Abasic (AP) sites are ubiquitous DNA lesions that are mutagenic and cytotoxic. Some BER glycosylases are bifunctional and cleave DNA at a transiently formed AP site via a lyase process (Stivers et al, 2003). In other instances, AP sites are produced as metastable intermediates. AP sites also are generated via spontaneous hydrolysis of native and damaged nucleotides.
DNA polymerase β (Pol β), a bifunctional enzyme that contains an 8 kDa lyase active site separate from its polymerase active site (Matsumoto and Kim, 1995; Matsumoto et al., 1998; Prasad et al., 1998), plays an integral role in BER by excising the remnant of an AP site following 5′-incision by apurinic endonuclease I (Ape1), and subsequently filling in the single nucleotide gap (Scheme 1).

Pol β excises the 5′-phosphorylated 2-deoxyribose (dRP) produced upon Ape1 incision of DNA containing an AP site (Scheme 2). Lys72 is the primary amine responsible for Schiff base formation, although the enzyme retains some lyase activity when this amino acid is mutated (Deterding et al., 2000; Prasad et al., 2005; Prasad et al., 1998; Feng et al., 1998). Lys84, which also is present in the lyase active site, is postulated to substitute for Lys72 in the mutated enzyme, albeit with much lower efficiency. Following Schiff base formation, dRP elimination leaves a single nucleotide gap that contains the appropriate end groups for DNA synthesis (by Pol β) and ligation to complete repair (Scheme 1, above).

Part of the attraction of Pol β as a potential therapeutic target is that it is over expressed in a variety of cancer cells (Husain et al., 1999; Starcevic et al., 2004; Barakat et al., 2012). In addition, Pol β variants are found in a large percentage of tumors (Donigan et al., 2012a; Nemec et al., 2012; Donigan et al., 2012b). Some of the variants exhibit reduced activity and may contribute to tumorigenesis by decreasing genomic stability. In addition, Pol β's vitality to genome integrity is manifested by the observation that cells lacking both alleles of the gene for this enzyme are embryonic lethal and knocking down Pol β activity sensitizes cells to DNA damaging agents (Horton et al., 2008). Consequently, Pol β has attracted interest as a target for antitumor therapy. Inhibiting Pol β potentiates the cytotoxic effects of DNA damaging agents and can be cytotoxic in its own right.
Natural and unnatural products have been tested as inhibitors of Pol β and the related enzyme, Pol λ, which is believed to act as a back up for Pol β in BER (Braithwaite et al., 2010; Braithwaite et al., 2005; Gao et al., 2008; Nakamura et al., 2007; Strittmatter et al., 2011; Wilson et al., 2010). Some of these molecules are believed to target the lyase domain. There is an interaction between Pol β and a DNA lesion, dioxobutane (DOB), which is produced by a family of potent cytotoxic antitumor antibiotics following C5′-hydrogen atom abstraction (Pitié and Pratviel, 2010; Goldberg, 1991). DOB efficiently inactivates Pol β (and Pol λ) (Guan and Greenberg, 2010; Guan et al., 2010; Stevens et al., 2013). Radiolabeling experiments, liquid chromatography, and mass spectral analyses of protease digests indicate that the 1,4-dicarbonyl inactivates Pol β in two ways (Scheme 3).

DOB forms a stable lactam following condensation with Lys72 or Lys84, elimination, and dehydration. The lesion also forms a stable adduct without undergoing DNA cleavage. The pC4-AP that is produced upon Ape1 incision of C4-AP (structures below) also contains a 1,4-dicarbonyl and inactivates Pol β and Pol λ (Stevens et al., 2013; Jacobs et al., 2011).
